Analysis of the age of Panax ginseng based on telomere length and telomerase activity

Abstract

Ginseng, which is the root of Panax ginseng (Araliaceae), has been used in Oriental medicine as a stimulant and dietary supplement for more than 7,000 years. Older ginseng plants are substantially more medically potent, but ginseng age can be simulated using unscrupulous cultivation practices. Telomeres progressively shorten with each cell division until they reach a critical length, at which point cells enter replicative senescence. However, in some cells, telomerase maintains telomere length. In this study, to determine whether telomere length reflects ginseng age and which tissue is best for such an analysis, we examined telomerase activity in the main roots, leaves, stems, secondary roots and seeds of ginseng plants of known age. Telomere length in the main root (approximately 1 cm below the rhizome) was found to be the best indicator of age. Telomeric terminal restriction fragment (TRF) lengths, which are indicators of telomere length, were determined for the main roots of plants of different ages through Southern hybridization analysis. Telomere length was shown to be positively correlated with plant age, and a simple mathematical model was formulated to describe the relationship between telomere length and age for P. ginseng.

Figure 1. In Situ Localization of TTTAGGG Telomeric Motifs on P. ginseng Chromosomes.

The the digoxigenin–dUTP nick tag sequence (CCCTAAA)₅ telomeric probe was hybridized with adventitious root of P. ginseng metaphase chromosomes and counterstained with propidium iodide.

Figure 2. The growth rings in the ginseng root of 1 ~ 6 years.

(A), (B), (C): 1 year ginseng root, 2 year ginseng root, 3 year ginseng root, Bar = 1000 μm; (D), (E), (F): 4 year ginseng root, 5 year ginseng root, 6 year ginseng root, Bar = 500 μm.

Figure 3. Developmental Regulation of Telomerase Expression in 5 years P. ginseng.

Telomerase activities in various tissues and different stages of plant development were assayed by TRAP, using 47F as the forward primer and PTelC3 as reverse primers.Lane1: tap root; Lane2: leaves; Lane3: stems; Lane4: root tips; Lane5: seeds.

Figure 4. Anatomical observation of 5 years P. ginseng by tangential cryosectioning.

Aseries of 400-um-thick tangential cryosections (A) ~ (J) was taken for each sample: tissues at different stages were isolated by tangential cryosectioning; (B): Telomerase activities in various tissues and different stages of plant development were assayed by TRAP, using 47F as the forward primer and PTelC3 as reverse primers; (C): Densitometric quantization revealed higher relative telomerase activity (relative units) in cambiums.

Figure 5. Southern hybridization images used for measurement and quantization of TRF length.

Lane M: DNA Molecular Weight Marker III, Digoxigenin-labeled (Roche). Numbers 2, 3, 4, 5, 6, 8 means different years of P. gensing samples collected from the city of Ji'an, Jilin province, China. B: Data fitting results and the trend of variation of TRF length with different ages. Overall, average TRF length increased with ages in main root (The following 1 cm of “ginseng lutou”).